Mucus plugging is a common pathological feature of allergic airway diseases, such as asthma, and causes small airways obstruction, subsegmental lung collapse, and occasionally precipitates respiratory failure. Effective therapies for mucus plugging in Th2 type inflammatory airway diseases are lacking because of an inadequate understanding of the basic pathophysiology that drives the formation of thick dehydrated mucus. Contrary to the current paradigm that Th2 type cytokines promote airway liquid secretion, we have found that the Th2 type cytokine IL-13 promotes airway surface liquid (ASL) hyperabsorption leading to profound impairments in ciliary function and mucociliary transport in primary cultures of human bronchial epithelial cells. Interestingly, Th2 cytokine mediated ASL absorption occurs via non- electrogenic ion exchange, a mechanism of bulk fluid absorption never previously attributed to airway tissue. Thus, airway surface dehydration presents a novel mechanistic explanation for the development of mucus dysfunction in subgroups of asthma patients. More importantly, restoration of mucosal hydration in asthma patients with intense Th2 inflammation presents a compelling therapeutic target to mitigate mucus obstruction. To address the hypothesis that IL-13 causes ASL absorption though coupled Na/H and Cl/HCO exchange and thus promotes pathological mucus dysfunction during Th2 inflammation, two complementary aims are proposed. The first aim will determine the ion conductance and signaling pathways that mediate ASL volume absorption by Th2 cytokines. In primary cultures of human bronchial epithelium and relevant heterologous expression systems, the following hypotheses will be tested (i) NHE3 and pendrin activities increase following IL-13 exposure as assessed by monitoring the rate of intracellular and extracellular pH change, (ii) NHE3 or pendrin gene knockdown prevents IL-13 dependent ASL hyperabsorption, and (iii) NHE3 transcription increases following IL-13 binding to the IL-4R / IL-13R1 heterodimeric receptor and activation of the STAT3 signaling pathway. Complementary to these in vitro experiments, the clinical 2nd aim will determine whether asthmatics with biological markers consistent with Th2 driven inflammation exhibit excessive ASL absorption, impaired mucociliary clearance, and increased NHE3/pendrin expression. Using a novel nuclear medicine technique that simultaneously measures in vivo airway fluid absorption and mucociliary clearance, airway clearance rates of patients with either Th2 low or high inflammatory profiles will be compared. Tissue samples will be collected by bronchoscopy and used to correlate NHE3 and pendrin expression with markers of Th2 inflammation and with rates of airway liquid absorption. Confirmation that Th2 inflammation promotes excessive ASL absorption and causes mucus obstruction, via NHE3 and pedrin, would establish a new conceptual framework and therapeutic avenue to manage mucus dysfunction in patients with asthma and other allergic airway diseases. PUBLIC HEALTH RELEVANCE: Mucus obstruction of the airway is a common but poorly understood feature of asthma and other allergic airway diseases. This project is based on the central hypothesis that mucus dysfunction occurs in these diseases because of airway surface dehydration, which impairs the primary innate mucociliary clearance mechanism from the lung. The information obtained from this research will provide insight into the pathogenesis of mucus plugging in allergic airway disease and may lead to innovative medical therapies.